Chemical Bulletin of “Politehnica” University of Timisoara, ROMANIA
Series of Chemistry and Environmental Engineering
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
Volume 57(71), 1, 2012
Headspace Solid Phase Micro Extraction and GC/MS Analysis
of the Volatile Components in Seed and Cake
of Azadirachta indica A. juss.
S. Shivashankar*, T.K. Roy* and P.N. Krishna Moorthy**
*
Division of Plant Physiology and Biochemistry, Indian Institute of Horticultural Research, Bangalore-560089, India
e-mail: siva@iihr.ernet.in
**
Division of Entomology and Nematology, Indian Institute of Horticultural Research, Bangalore-560089, India
e-mail: pnkm@iihr.ernet.in
Abstract: The present investigation was carried out to identify the components present in the headspace volatiles of neem
seed and cake as these have been found to exert insect repellant action on many pests of horticultural crops. The
composition of volatiles emitted by neem seed (Azadirachta indica L.) and its cake obtained after extraction of oil was
analyzed by headspace solid-phase micro extraction (HS-SPME) combined with gas chromatography and gas
chromatography- mass spectrometry (GC-MS). Major differences were found in the volatile composition of seed and cake.
A total of 71 and 101 volatile compounds representing 83.47 % and 85.62 % of the total quantum of volatiles present in
seed and cake, respectively, could be identified. These compounds were found distributed in several chemical classes
namely, alcohols, carbonyl compounds (ketones, aldehydes, and esters), fatty acids, terpenes, nitrogenous and sulphur
compounds among others. Of the various classes of compounds identified, fatty acids constituted the major group
accounting for 24.82 % and 16.97 % of the total volatile concentration followed by aldehydes and ketones representing
26.49 % and 17.53 % and sulfur compounds contributing 12.94 % and 9.32 % of seed and cake volatiles, respectively. The
most abundant volatile components were (Z)-9,7-octadecadienal and palmitic acid, a medium chain fatty acid comprising
25.47 % and 14.97 % of the total in seed and cake respectively.
Keywords: Azadirachta indica, neem seed and cake, neem volatiles , SPME, GC-MS.
major component, di-n-propyl disulfide exhibited larvicidal
properties. Subsequently, the presence of certain volatile
compounds in neem leaf having fungicidal properties were
reported [8]. Neem seed volatiles extracted by organic
solvents were also found to be effective against the eggs
and adults of pulse beetle, Callosobruchus maculatus [9].
Keeping in view, the ecological safety and possible
commercialization potential of neem volatiles for insect
pest control in horticultural crops, the present work was
carried out with the aim of separating and identifying the
entire range of volatile compounds emitted by seed and
cake using GC-GCMS. Previous studies on the head space
volatiles of other seeds reported advantages of using SPME
[10, 11] to avoid interferences from nonvolatile matrix
components. Based on these reports, headspace sampling
was employed for the extraction of volatiles in the present
study.
1. Introduction
The
neem
tree,
Azadirachta
indica
A.Juss.(Meliaceae), is indigenous to India and Southeast
Asia. Extracts of neem fruit, seeds, seed kernels, twigs,
stem bark and root bark have been shown to possess insect
antifeedant, insecticidal, insect growth disrupting,
nematicidal, fungicidal, bactericidal, anti-inflammatory,
antitumour, immuno-stimulating and many other biological
activities [1]. In India, neem cake, a byproduct produced
during extraction of oil from neem seed, is used as a
biopesticide. The use of neem cake as an amendment to
agricultural soils to control plant parasitic nematodes under
both greenhouse and field conditions is a well known
practice in India [2, 3, 4]. Field studies conducted by us
showed that the volatiles emitted from neem cake were
effective in controlling diamondback moth (DBM),
Plutella xylostella of cabbage, a notorious insect pest
known to develop resistance against most of the
commercially available pesticides and other insects
affecting vegetable crops [5]. Although several reviews
have been published on the chemical constituents of neem
seed [6], the chemical profile of volatiles emitted by both
neem seed and cake have not received much attention
excepting a report [7] on the volatile organosulfur
compounds of neem seed in which it was shown that the
2. Experimental
2.1. Plant material
Neem trees growing near the experimental farm of the
institute were used for the study. The experimental site was
characterized by a mild tropical weather throughout the
year with a mean maximum temperature of 28-30 0C and a
1
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
Volume 57(71), 1, 2012
minimum of 17-20 0C during the fruit development phase.
Mature neem fruits were harvested during the month of
August and ripened under ambient conditions. Seeds from
ripe fruits were separated from the pulp, washed with water
and dried in the shade for a week. The kernel obtained after
decorticating the seeds was crushed in a hydraulic press at
a high pressure in a single step to extract the oil. The
resulting neem cake containing 8 % oil was ground into a
fine powder and used for the experiment. Neem seed
powder was prepared by finely pulverizing the seeds in a
tissue grinder immediately before use.
0.25 mm ID with 0.25 µm film thickness) by applying the
same temperature program as described above for GC-FID
analysis. Mass detector conditions were: EI-mode at
70 eV, injector temperature, 250 °C; ion sourcetemperature, 230 °C; trap temperature, 220 °C; transfer line
temperature 250 °C and full scan range, 50–450 amu. The
carrier gas was helium at a flow rate of 1 ml.min-1.
2.5. Identification of components
Volatile compounds were identified by comparing the
mass spectra with the data system libraries (Wiley and
NIST-2007) and by Kovats indices (KI) computed in
accordance with modified Kovats method [13,14 ] using a
homologous series of n-alkanes (C5 to C32). The Kovats
index, for the unknown was calculated from the formula:
2.2. Isolation of volatile components
Extraction of head space volatiles of neem seed and
cake was performed as described earlier [12] with some
modifications. The SPME fibre coated with carboxanpolydimethylsiloxane-divinylbenzene
(50/30µm,
CAR/PDMS/DVB) (Supelco, Bellefonte, PA, USA) was
used for the analysis, due to its high sensitivity for aroma
compounds and excellent reproducibility. 50 g each of seed
and cake were homogenized with 100 mL double distilled
water using a commercial blender. The slurry was
transferred to a 250 ml conical flask to which 5 g of NaCl
was added. Subsequently, the flask was sealed with a
teflon-lined septum and the samples were kept stirred at
37±1°C. After 20 min of equilibration between the solution
and the headspace, the fibre was exposed to the headspace
of the sealed flask for 60 min. Prior to sampling, the fibre
was preconditioned for 1hr at 260°C in the GC injection
port as per the manufacturer’s instructions.
KI = 100[n +(N - n) X {log tr(unknown) – log tr(n)}/{log
tr(N) – log tr(n)}]
where (n) is the number of carbon atoms in the shorter
chain alkane, N is the number of carbon atoms in the longer
chain alkane, (tr) is the adjusted retention times for
unknown compounds, tr (n) , shorter-chain and tr (N),
longer-chain alkanes.
The total volatile production was estimated as the sum
of all GC-FID peak areas in the chromatogram and
individual compounds were quantified as relative percent
area. All data are means of three independent
determinations expressed as percentage of total peak area.
2.3. Gas chromatography
3. Results and Discussion
GC / FID analysis was carried out using a Varian-3800
Gas chromatograph system with SPME sleeve adapted to
injector on a VF-5 column (Varian, USA), 30 m x 0.25 mm
i.d, and 0.25 µm film thickness. The carrier gas was helium
at a flow rate of 1ml min-1; injector temperature 250 °C and
detector temperature 270°C. The temperature program for
column was as follows: The initial oven temperature was
50 °C for 2 min , increased by 3 °C /min up to 200 °C,
held for 3 min, increased further at 10 °C/min up to 220 °C
and maintained constant for 8 min. For desorption, the
SPME device was introduced in the injector port for
chromatographic analysis and remained in the inlet for 12
min. Initially injection mode was splitless followed by split
mode (1:20) after 1.5 minutes. For the qualitative
identification of volatile substances and computation of
retention time and index, the following standards, ethyl
acetate, propanol, isobutanol, hexanol, 1-octene-3-ol and
eugenol were co-chromatographed.
2.4. Gas
(GCMS)
chromatography/mass
A total of 71 and 101 volatile compounds could be
separated and identified from the headspace samples of
neem seed and cake powder respectively (Table 1). Of
these, 57 compounds were common to both. The number of
sulfur compounds was 20 in both neem seed and cake
accounting for 12.94 % and 9.32 % of the total amount of
volatiles respectively. The number of hydrocarbon
compounds in cake was higher at 22 as against 17 in seed
but their proportion increased from 2.74 % in seed to 18.33
% in cake. The relative proportion of oxides, esters and
miscellaneous compounds in cake increased appreciably as
compared to those in seed. On the other hand, the relative
contribution of sulfur compounds, acids, aldehydes and
ketones showed a marked fall in cake as compared to seed
(Table 1). There were no marked differences in either the
number or proportion of nitrogen compounds present in
seed and cake. Of the various groups of compounds
identified, aldehydes and ketones constituted the major part
(26.49 %) of seed volatiles followed by acids accounting
for 24.82 %. On the contrary, the proportion of acids was
higher in cake volatiles (18.54 %) followed by aldehydes
and ketones (17.53 %). Palmitic acid was the most
abundant fatty acid compound in both seed (24.34 %) and
cake (14.97 %) followed by (Z)-9, 17-octadecadienal
which constituted 25.47 % and 10.57 % of seed and cake
respectively. Besides these two compounds, other major
spectrometry
GC-MS analysis was performed on Varian-3800 gas
chromatograph coupled with Varian 4000 GC-MS/MS
mass selective detector. Volatile compounds were
separated on VF-5MS (Varian, USA) column ( 30 m x
2
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
Volume 57(71), 1, 2012
volatiles found in seed were, 5-chloro-1H-indole (10.27%),
2-methoxythiophene (3.44 %), 4-benzylfuro [3, 2-c]
pyridine (3.16 %), 2-furylmethylsulfide (1.57 %) and 2,5dimethylthiophene (3.06 %). Similarly, the major volatile
compounds found in neem cake included, 3, 5-diethyl-1, 2,
4-trithiolane (2.25 %), 1, 5-dithiaspiro [5,7] tridecan-7-ol
(1.79 %), 4-benzylfuro(3, 2-c ) pyridine (5.74 %), 5-chloro1H-indole (2.73 %), (E)-3-(2-Quinolyl) propenenitrile
(2.80 %), 3-furaldehyde (2.29 %), 1-methylene-1H-indene
(6.59%),4-hexyl-2,5-dihydro-2,5-dioxo-3-furanylaceticacid
(1.28 %), 1-undecyne (1.28 %), zingiberene (1.49 %) and
bisabolene (2.01 %).
TABLE 1. SPME headspace analysis of volatile compounds of neem seed and cake
Sl.
No.
Class and compound
34
35
36
37
38
39
40
41
42
43
44
45
Sulfur compounds
Carbon disulfide
Methyl-thiirane
(1E)-1-(Methylsulfanyl)-1-propene
Methyl-3-thiophene
Thiophene, 2-methoxyThiophene, 2,5-dimethylThiophene,3,4-dimethyl2-Furylmethylsulfide
2,3-Butanedithiol
(1E)-1-(Methyldisulfanyl)-1-propene
1,2-Dithiacyclopentane
Isothiazole, 4,5-dimethyl1,3-Dithiane
2-Ethyl[1,3]dithiane
2-Thiaadamantan-4-ol
3,5-Dimethyl-2-(methylsulfanyl)thiophene
1-Methoxy-4-(methylthio) benzene
5-methyl-4,7-dithiadeca-1,9-diene
Propenyl propyl trisulfide
1,2,4-Trithiolane, 3,5-diethyl[4-(Methylsulfanyl)phenyl]methanol
2,3-Thiophenedicarboxylic acid
2,3,4,5-Tetrathiabicyclo[4,4,0]decane
1,5-Dithiaspiro[5.7]tridecan-7-ol
Nitrogen compounds
3,4-Dihydro-2H-pyran
1H-Pyrrole-2-carboxaldehyde
1H-Pyrazole, 3,5-dimethyl4-benzyl furo[3.2-c] pyridine
2-Ethyl-3,5,6-trimethylpyrazine
1H-Indole, 5-chloro7-Chloro-s-trizolo(pyrimidine-5(1H)-one
2-Propenenitrile, 3-(2-quinolinyl)-, (E)(9Z)-octadec-9-enenitrile
Alcohol Compounds
4-Methyl-1-penten-3-ol
3-Hexyn-2-ol
3-Hexen-1-ol, (Z)5-Hexen-1-ol
2-Cyclohexen-1-ol
Cyclohexanol, 4-methyl6-Methyl-6-hepten-4-yn-3-ol
(2E,6E)-2,6-Octadiene-4,5-diol
α-Methyl-1,4-benzenedimethanol
(E)-Isoeugenol
Nerolidol
Spathulenol
46
Ledol
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
Kovats Index (KI)
Calculated
Reported
569
652
684
788
878
882
889
902
910
922
985
988
1031
1076
1222
1230
1248
1256
1311
1359
1419
1618
1682
1961
568
650
678
794
NA
882
888
894
902
922
978
974
1027
1063
NA
NA
NA
NA
1318
NA
NA
NA
NA
NA
668
1012
1024
1112
1139
1361
1669
1686
2126
NA
988
1014
N.A
1120
N.A
N.A
N.A
NA
708
796
853
858
890
926
972
1186
1422
1455
1572
1581
704
797
851
NA
887
929
NA
NA
1411
1459
1564
1576
1621
1608
3
*Area (%)
Neem Seed
Neem Cake
0.064
0.404
0.297
0.096
3.441
3.059
0.659
1.572
0.085
0.149
0.340
0.202
0.836
0.372
0.276
0.085
0.096
0.127
0.542
0.234
3.165
10.271
0.329
0.276
0.446
0.064
0.096
0.202
0.096
0.085
0.096
0.149
0.123
0.113
Detector used
1.796
FID
FID
FID
FID
MS
FID
FID
FID
FID,MS
FID,MS
FID
FID
FID,MS
MS
MS
MS
MS
MS
FID
MS
MS
MS
MS
MS
0.062
0.667
0.062
5.738
0.092
2.731
0.749
2.803
0.626
MS
FID,MS
FID,MS
MS
MS
MS
MS
MS
MS
0.082
0.062
0.092
0.113
0.082
0.092
0.195
0.729
1.006
0.842
FID
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
MS
MS
FID,MS
FID,MS
FID,MS
FID,MS
0.390
FID,MS
0.082
0.390
0.431
0.113
0.359
0.082
0.092
0.216
0.082
0.195
0.113
0.257
0.955
0.534
2.248
0.267
0.873
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
47
48
49
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
1-(1-Naphthyl)-2-propanol
1,2-Benzenediol, 3,5-bis (1,1dimethylethyl)(6Z,9Z)-6,9-Pentadecadien-1-ol
Aldehydes and ketones
Furfural
3-Furaldehyde
4,6-Heptadiyn-3-one
1-Methyl-1-cyclopenten-3-one
2,4-Hexadienal, (E,E)Octenal
1-(1H-pyrrol-2-yl)-Ethanone
(2E,4E)-2,4-Nonadienal
4-(1,2-Dimethyl-cyclopent-2-enyl)-butan--22-one
Ethanone,
2-hydroxy-1-phenyl10-Undecenal
Dihydro-β-ionone
1H-2-Benzopyran-1-one, 3,4-dihydro-8hydroxy-3-methylBenzophenone
Isolongifolen-5-one
1,4-cis-1,7-cis-Acorenone
(E,E)-10, 12-Hexadecadienal
(Z)-9,17-Octadecadienal
Acids
2-Cyclopentene-1-carboxylic acid, 1-methylDodecanoic acid
Tetradecanoic acid
Pentadecanoic acid
4-Hexyl-2,5-dioxo-2,5-dihydro-3furanyl)acetic
acid
Palmitic acid
Esters
Phenethyl acetate
Methyl (6E)-3,7,11-trimethyl-6,10dodecadienoate
Benzyl
Benzoate
Methyl hexadecadienoate
Methyl hexadecanoate
Ethyl palmitate
Methyl (7E,10E)-7,10-octadecadienoate
Methyl linolenate
Ethyl linoleate
Hydrocarbons
Toluene
1,3,7-Octatrien-5-yne
4-Nonene, 5-methyl1-Methylene-1H-indene
1-Undecyne
Naphthalene
α-Longipinene
Ylangene
β-Elemene
Copaene
α-Gurjunene
Elixene
Acoradiene
β-Eudesmene
α-Calacorene
α-Muurolene
δ-Cadinene
Valencene
Zingiberene
α-Amorphene
103
Bisabolene
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
Volume 57(71), 1, 2012
1712
1788
1792
NA
NA
1771
842
846
868
885
909
1059
1065
1235
1265
1277
1288
1433
1532
1628
1678
1698
1702
2018
831
832
868
887
908
1060
1063
1218
1240
NA
1278
1432
1514
1612
NA
NA
NA
1997
1085
1569
1772
1822
2118
1978
1073
1566
1760
1806
NA
1961
1251
1698
1695
1869
1878
1991
2101
2108
2165
1260
1685
1693
1881
1890
1991
2093
2098
2155
764
828
1015
1098
1109
1186
1371
1381
1386
1388
1436
1449
1479
1488
1493
1498
1522
1512
1515
1518
759
822
1002
1095
1096
1179
1353
1371
1391
1376
1403
1431
1471
1483
NA
1495
1536
1499
1500
NA
0.085
0.191
0.096
0.159
0.191
0.159
1521
1507
0.191
4
0.117
0.096
0.113
0.267
0.082
MS
MS
FID,MS
0.062
2.290
0.636
0.123
10.574
FID
FID
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
MS
FID,MS
FID,MS
FID,MS
FID,MS
MS
MS
MS
FID,MS
0.287
0.770
0.903
0.327
1.283
14.970
FID,MS
FID,MS
FID,MS
FID,MS
MS
FID,MS
0.062
0.709
FID,MS
MS
FID,MS
FID,MS
FID,MS
FID,MS
MS
FID,MS
FID,MS
0.276
0.127
0.191
0.149
0.082
0.216
0.113
0.955
0.226
0.082
0.832
0.708
0.734
0.096
0.117
0.064
25.469
0.191
0.127
0.159
24.340
0.266
0.085
0.085
0.096
0.191
0.085
0.159
0.096
0.266
0.223
0.113
0.595
0.873
0.226
0.154
0.257
0.062
0.051
6.589
1.282
0.431
0.283
0.915
0.749
0.606
0.287
0.133
0.400
0.082
1.039
1.486
0.216
FID
FID
FID
FID
FID
FID
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
FID,MS
MS
FID,MS
FID,MS
FID
FID,MS
MS
2.013
FID,MS
0.185
0.117
0.127
0.329
0.212
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
Volume 57(71), 1, 2012
104
105
106
1-Pentadecyne
1526
Juniper camphor
1688
Podocarpa-8,11,13-triene, 13-isopropyl2078
Oxides
107
1,2-Dimethoxybenzene
1150
108
2,3-Dimethoxytoluene
1169
109
1,4-Dimethoxybenzene
1172
110
Caryophyllene oxide
1579
Others
111
2,5-Dimethyl furan
718
112
2,4-Dimethyl furan
729
113
2-Ethenylfuran
765
114
2-Pentylfuran
996
115
2,6-di-tert-Butyl-p-benzoquinone
1482
Total identified constituents
*Data are expressed as percentage of the total peak area
1510
1675
2054
Earlier attempts in our laboratory to extract the volatile
compounds of neem cake by steam distillation followed by
partition with organic solvents and concentration under
vacuum gave extracts in which many volatile constituents
were identified [15]. However, based on the previous
reports [16, 17] in which it was shown that certain
compounds added to neem leaf could not be detected after
boiling of neem leaves for extraction of volatiles, it was
suspected that many volatile components might have either
escaped due to their high volatility or had undergone
changes in their structure and activity due to the extreme
conditions of temperature and pressure employed in the
study. Therefore, SPME was used in the present analysis as
it facilitated rapid sample preparation from a small amount
of sample and with little or no change in structure and
composition in comparison to cold press or steam
distillation [18, 19]. GC MS analysis of SPME samples
gave 20 organosulfur compounds from seed and 21 from
cake which were chemically distinct from those reported
earlier [7]. It must be noted that the above study reported
the presence of 22 volatile organosulfur compounds from
the headspace samples of neem seed collected by purging
with a continuous stream of nitrogen gas followed by
extraction using diethyl ether solvent. It was apparent
therefore, that the differences in the nature and composition
of separated volatiles were the result of the differences in
the extraction protocols. On the contrary, the differences in
the composition of sulfur compounds of neem cake
samples from those reported by Krist et al.,[20] could have
arisen
from the enzymatic degradation of sulphurcontaining compounds of seed .
The two most abundant individual components of seed
were (Z)-9,7-octadecadienal and palmitic acid, a medium
chain fatty acid comprising 25.47% and 24.34% of the total
respectively. On the other hand, palmitic acid was the most
abundant individual component of cake (14.97%) followed
by (Z)-9,7-octadecadienal forming 10.57% of the total.
Incidentally, a recent study [21] also reported that the most
abundant fatty acid present in Azadirachta indica and
Azadirachta siamensis was palmitic acid. Palmitic acid has
been reported to be active in Aedis aegypti biting deterrent
assays [22]. The levels of acids, aldehydes and ketones
were lower in cake as compared to seed while there was an
increase in the levels of esters in cake. This could perhaps
0.064
0.257
0.780
0.226
FID
FID,MS
FID,MS
1150
1172
1186
1582
0.113
0.113
0.257
0.287
FID
FID,MS
FID
FID,MS
695
708
761
991
1462
0.062
0.051
0.060
0.154
0.287
85.617
FID
FID
FID
FID
MS
0.074
0.085
0.117
83.475
be due to the action of seed enzymes on the cellular
substrates which are released due to cellular damage
occurring during the extraction of oil from seed. The
marked increase in the levels of oxides and miscellaneous
compounds in neem cake indicated increased oxidation and
hydrolysis of seed components by hydrolytic and oxidative
enzymes released due to tissue damage during cake
formation.
In an earlier study, we had observed that the volatiles
from neem cake exhibited insect repellant activity on the
diamondback moth (Plutella xylostella) of cabbage
(Unpublished). Based on the data obtained in the present
study, it should be possible to identify the components of
volatiles of neem cake responsible for the insect repellant
activity. It has been reported that volatile compounds
emitted from neem leaf exert significant inhibitory effects
on fungal growth and aflatoxin production in aflatoxigenic
Aspergillus parasiticus cultures [8]. Short term exposure to
neem volatiles was found to impair the gonotrophic cycle
and suppress oviposition of female Anopheles while long
term exposure impaired vitallogenosis irreversibly [23]. It
is noteworthy that diamondback moths exposed to a
mixture of neem compounds for 35 successive generations
did not show any signs of incipient resistance [24].
Volatiles from neem seed have also been reported to be
effective against stored grain pests [9, 25, 26]. Several
studies have established the distinctive activity and
function of neem components in either altering insect
behavior or life processes in such a subtle manner that the
insect would be unable to feed, breed, metamorphose or
cause damage to plants [24]. Based on these findings, it is
imperative that the volatiles of neem seed and cake
reported in this paper could have potential application for
management of insect pests of crops and mosquito control.
Neem seed and cake being natural products with a long
recorded history of use in agriculture in India would be
ideal candidates for successful mosquito control and ecofriendly management of insect pests of crops.
Supplemental information on the MS spectra analysis
I= 878, [M+] 114, m/z 114(100) 99(97)55(55) 71(42). I=
1076, [M+] 146, m/z 146(100)72(65.5)118(15). I= 1222,
5
Chem. Bull. "POLITEHNICA" Univ. (Timisoara)
Volume 57(71), 1, 2012
[M+] 170, m/z 70(100) 79(56)99(41)142 (35)67(20). I=
1230, [M] 158, m/z 143(100) 158(90) 99(19) 59(11). I=
1248, [M+] 154, m/z 139(100) 154(97.5) 111(12)
77(8)95(7).
I=
1256,
[M+]
188,
m/z
73(38)59(33)115(18)147(17)101(11)188(20).
I= 1359,
[M+] 180, m/z 74(79) 180(26) 106(18)59(18)116(12). I=
1419, [M+] 154, m/z 154(100)107(35) 79(34) 137(29)
109(24)125(20).
I= 1618, [M+] 172, m/z
111(100)
128(62) 172(32) 83(21) 155(14) 82(13) 57(11). I= 1682,
[M+] 210, m/z 210(100) 81(60) 146(14) 113(10) 82(7)
145(4)67(3). I= 1961, [M+] 232, m/z 145(100) 125(80)
106(52) 119(28) 232(27) 95(27) 55(23)132(17)73(16). I=
668, [M+] 84, m/z 55(100)84(51)56(32)54(26)83(26). I=
1112, [M+] 209, m/z 207 (100) 194(20) 180(6) 134(5). I=
1139,
[M+]
150,
m/z
49(100)
150(83)53(25)54(21)122(18)67(14)56(13). I= 1361, [M+]
151,
m/z
151(100)
89(26)116
(19)124(11)75(9)63(7)115(5). I= 1669, [M+] 170, m/z
170(100) 172(35) 107(15) 129(10) 142(8). I= 1686, [M+]
180, m/z 180(100)154(24)179(24)181(14)155(4). I= 2126,
[M+]
263,
m/z
55(100)
69(50)122(48)136(40)83(35)97(28)150(20)220(15)234(8)2
124,
m/z
95(100)
63(7).
I=
972,
[M+]
67(25)65(23)109(15)51(13)91(9)77(8)66(7). I= 1186, [M+]
142,
m/z
71(100)72(10)
53(10)
57(6)
69(5)55(4)125(4)70(3).
I= 1712, [M+] 186, m/z 42
(100)141(59)186(17)115(16.8)
143(12)
139(6)
222,
m/z
63(4)128(3).
I=
1788,
[M+]
207(100)222(20)57(19)208(15)179(5)91(5)77(4)191(4). I=
1277,
[M+]
136,
m/z
105(100)77(60)51(16)106(8)136(6)78(5)50(4). I= 1678,
[M+] 218, m/z 175(100) 218(69)147(68)119(66)105(43)162
(35)133(34)91(29)134(28).
I= 1702, [M+] 236, m/z
55(100) 67(80)81(60)96(35)236(25). I= 2018, [M+] 264,
m/z 67(100)81(72) 55(58) 95(45) 264(20).
I= 2118,
[M+] 240, m/z 126(100)98(21)140(16)95(12). I= 1978,
[M+]
256,
m/z
73(99)60(84)
57(63)
55(62)
69(31)71(28)61(22).
I= 1698, [M+]
252,
m/z
69(100)109(89)209(48)123(37)95(22)81(18)67(17)
55(14)177(14). I= 2101, [M+] 294, m/z 67(100)81(
90)55(70)95(67)82(54)79(48)96(45)68(43). I= 1493, [M+]
200,
m/z
157(100)142(52)141(35)156(24)158(14)115(14)200(11)14
3(9)128(9).
I=
1518,
[M+]
204,
m/z
61(100)105(72)119(56)91(52)93(46)79(41)133(31)77(30)2
04(28).
I=
1433,
[M+]
194,
m/z
121(100)161(43)136(36)123(36)93(33)95(27)81(25)119(2
3).
I=
1698,
[M+]
220,
m/z
82(100)109(56)135(45)123(40)220(36)55(31)136(28)121(
26.5)|93
(25).
I=
1482,
[M+]
220,
m/z
177(100)220(73)67(37)135(33)57(33)149(30)
205(28)163(24).
found effective in controlling insect pests of horticultural
crops, it would be worthwhile extending the study further
to identify the specific components which are effective
against insect pests. It is likely that detailed studies on the
structure and mode of action of the biologically active
volatile components would open up a new generation of
ideal pesticides for the future.
ACKNOWLEDGEMENTS
The authors wish to thank the Director of the institute
for providing facilities for the study and Mr. S.S.A. Qazi
for the skilled assistance in the analysis of samples.
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4. Conclusion
The present work on separation and characterization of
a large number of volatiles from both neem seed and cake
has established that the neem seed cake is a better source of
volatiles than the seed per se. Since these volatiles were
Received: 16 April 2012
Accepted: 05 June 2012
6